Lecture Goals Microbiologic Sampling of the Environment

Microbiologic Sampling of the Environment

William A. Rutala, Ph.D., M.P.H., C.I.C.Director, Statewide Program for Infection Control and Epidemiology and Professor of Medicine, University

of North Carolina at Chapel Hill, NC, USA Former Director, Hospital Epidemiology, Occupational Health and Safety, UNC Health Care, Chapel Hill, NC

(1979‐2017)


•Lecture Goals•Microbiologic Sampling•Indications•Methods


• History• Pre‐1970, hospitals regularly cultured air and surfaces• By 1970, AHA advocated discontinuation because HAI not associated with levels of microbes in the air and surfaces; not cost‐effective

• In 1981, CDC recommended targeted sampling (eg, sterilizers and dialysis water)

Microbiologic Sampling of the EnvironmentCDC Guidelines for EIC, 2003

• Targeted microbiological sampling• Support of an investigation of an outbreak • Research• Monitor a potentially hazardous environmental condition• Quality assurance


• Will environmental sampling provide meaningful, interpretable data that help identify actual or potential contamination problems associated with a specific procedure or instrument• Should not be done if no plan for interpreting and acting on the results obtained• Is it justified on epidemiological grounds• No accepted criteria for defining surfaces or air as clean/safe in healthcare

Microbiologic Sampling of the EnvironmentInvestigation of an Outbreak

• When?•Environmental reservoirs or fomites are implicated epidemiologically in disease transmission•Plan for interpreting and acting on the results•Plan to link microorganisms from the environment with clinical isolates by molecular epidemiology

Microbiologic Sampling of the EnvironmentInvestigation of an OutbreakRutala et al. J. Thorac. Cardiovasc. Surg. 96:157‐161.

• Outbreak: two patients in CT‐ICU with symptomatic B. cepacia • Epidemiologic investigation: case‐control study revealed that both patients required an intra‐aortic balloon pump (IABP) for circulatory support• Microbiological investigation: water reservoir of IABP contained >105B. cepacia/ml

Microbiologic Sampling of the EnvironmentInvestigation of an OutbreakRutala et al. J. Thorac. Cardiovasc. Surg. 96:157‐161

• Microbiologic investigation: causative organism isolated from several components of the IABP and the hands of a nurse who manipulated the IABP’s buttons/switches.• Molecular epidemiology: similar plasmid profilefrom strains from the patients and the IABP.• Conclusion: transmission presumably occurred during manipulation of IV lines .

Microbiologic Sampling of the EnvironmentResearch

•When? Experimental methods that provide new information about the spread of HAIs• Example: Relation of the Inanimate Hospital Environment to Endemic Nosocomial Infection (NEJM 1982;302:1562).• Cultured air, surfaces, and fomites in old/new hospital and despite major differences in contamination (17% positive vs 5%), incidence of NI remained unchanged.

Objective and Design

• To determine if enhanced methods for terminal room disinfection decrease acquisition and infection due to multidrug‐resistant organisms (MDROs)

• Prospective, multicenter, cluster‐randomized, crossover trial to evaluate three strategies for enhanced terminal room disinfection• 9 hospitals• Randomization at level of hospital• 2x2 factorial design

Terminal Room Disinfection

No UV‐C UV‐C

Quat* A B

Bleach C D

*NOTE: Bleach always used in rooms of patients with suspected or confirmed C.

difficile

Definitions and Inclusion Criteria

Terminal Clean

Patient in “Seed Room”

Documented infection or colonization with MRSA VRE

C. difficileMDR‐Acinetobacter

“TARGET MDROs”

“Exposed”Patient

In room ≥ 24 hours

Exposure days = Time spent in “seed room”

Same organism as the patient in the “seed room” AND

Positive culture while in roomOR

Potential “Incident Case”

Positive culture after stay in room‐ 90 days (MRSA, VRE, MDRAB)‐ 28 days (C. difficile)

Enhanced Disinfection Leading to Reduction of Microbial Contamination and a Decrease in Patient Col/InfectionAnderson et al. Lancet 2017;289:805; Rutala et al. ICHE 2018

All enhanced disinfection technologies were significantly superior to Quat alone in reducing EIPs. Comparing the best strategy with the worst strategy (i.e., Quat vs Quat/UV) revealed that a reduction of 94% in EIP (60.8 vs 3.4) led to a 35% decrease in colonization/infection (2.3% vs 1.5%). Our data demonstrated that a decrease in room contamination was associated with a decrease in patient colonization/infection. First study which quantitatively described the entire pathway whereby improved disinfection decreases microbial contamination which in-turn reduced patient colonization/infection.

Microbiologic Sampling of the EnvironmentMonitor a Potentially Hazardous Environmental Condition

• When? Confirm the presence of a hazardous chemical/biological agent, and validate abatement of the hazard• Examples

• Detect bioaerosols (eg, ultrasonic cleaner, water fountain)• Detect agent of bioterrorism• Sample for IH (eg, sick building)

Microbiologic Sampling of the EnvironmentQuality Assurance

•When? To evaluate the effects of a change in infection control practice or ensure equipment/systems perform as expected• Air sampling during construction/renovation to qualitatively detect breaks in IC measures

• Only routine sampling recommended: biological monitoring of sterilizers, monthly cultures of water used in hemodialysis

• Endoscopes/AERs

Microbiologic Sampling of the EnvironmentAir Sampling

• General comments• Particles in a biological aerosol usually vary from <1 to >50 µm.• Particles consist of a single, unattached organism or clumps• Vegetative cells do not ordinarily survive long in air• Pathogens may settle on surfaces and become airborne again with sweeping, etc


• Air sampling for QA is problematic due to the lack of uniform air quality standards• The critical number of Aspergillus that poses a risk for neutropenic patients is not known• Results affected by factors (traffic, time of year)• Results need to be compared to other defined areas

Air Sampler Methods for Culturing AirJ Boyce 2012

Air SamplingJ Boyce 2012

• Settle plates can be expressed as number of bacteria in an area (e.g., patient room) for a specified time (e.g., 1 hour)• Liquid impingers can provide data on the number of particles/microbes per volume of air sampled (e.g., 100 CFU/20 ft3)• Volumetric Sieve samplers (e.g., Anderson) can size particles and sample specific volume (e.g., 20 CFU of respirable particles/20 ft3)


• Factors in Selecting an Air Sampling Device• Viability and type of organism• Skill required to operate sampler• Availability and cost of sampler• Availability of auxiliary equipment (vacuum pump)• Assumed concentration and particle size• Sensitivity of microorganisms to sampling• Compatibility with the selected method of analysis


• Impingement in liquids• Impaction on solid surfaces• Sedimentation• Filtration• Centrifugation• Electrostatic precipitation• Thermal precipitation


• Impingement in liquids‐collects (mo directed against a liquid [nutrient broth], conc over time)Ex. Water aerosols for Legionella • Impaction on solid surfaces (sieve)‐collects (mo deposited on agar), sizes, conc per unit volume of air (CFU/ft3). Ex. Aspergillus• Sedimentation (settle plates)‐mo settle on agar via gravity, conc over time (CFU/time)

Microbiologic Sampling of the EnvironmentWater Sampling

•When? Routine testing of water not indicated (except dialysis) but sampling in support of outbreak investigation can help determine IC measures• Use established methods (eg, sample water ASAP after collection, 100ml minimum, sterile collection equipment, neutralizers, recovery media and incubation temp [diluted peptone, 20oC], pour plates [high counts], membrane filtration‐0.2µ [low counts, larger volumes])• Filters are placed on agar plates and incubated for 48h

Filtering

Microbiologic Sampling of the EnvironmentSurface Sampling Methods

• Sample/Rinse‐use sterile wipe/sponge/wipe, media, qualitative/quantitative assays• Direct Immersion‐immerse in media, then assay• Containment‐interior surfaces of containers• RODAC (replicate organism detection and counting)‐sampling flat, nonabsorbent surfaces, direct assay

Methods for Culturing SurfacesCDC, 2003; Boyce, 2012

• Moistened swab (with template ideal)• Moistened swab and rinse (broth enrichment)• Moistened sponge and rinse• Moistened wipe and rinse• Direct Immersion• RODAC plates

• Irregular objects

• Irregular objects

• Large, flat surfaces

• Large, flat surfaces• Immerse in broth• Flat surfaces

Factors Affecting Organism RecoveryRawlinson et al. J Hosp Infect 2019;103:363

• Target organism and strain• Level of contamination• Wet/dry surface• Adsorption of cells• Pressure and contact time• Media• Pre‐wetting, enrichment

• Brand on contact plates• Cell injury/stressors• Size of surface sampled• Number of samples• Cost• Sensitivity• Difference in contamination

Moistened Swab with Direct PlatingBoyce, 2012

• Use moistened swab to sample surfaces• If defined area not sampled, results are semi‐quantitative• If defined area sampled using a template, results are quantitative (CFUs/cm2); preferable

•Moistening (wetting) agents include normal saline, broth media (neutralizers)• Swab is used to directly inoculate non‐selective or selective media, followed by incubation x 48h• Use for sampling irregular‐shaped objects

Moistened Swab with Direct PlatingBoyce, 2012

• Advantages• Easy to perform• Simple; can be used in many facilities with microbiology laboratory support• Provides information about general level of contamination or for specific pathogens

• Disadvantages• Least sensitive method for detecting or organisms on surfaces• Non‐standardized procedure makes comparison of studies difficult

RODAC PlatesBoyce, 2012

• Small petri plate filled with agar to provide convex surfaces• Agar plate is pressed against a flat surface, plate is incubated• Advantages: very easy to perform and standardized; results expressed as CFU/cm2 ( suggested clean 2.5 CFU/cm2 or 65CFU/Rodac); neutralizer available• Disadvantages: greater cost; sample small area per plate

Microbiologic Sampling of the EnvironmentSurface Sampling

• Used for research (potential reservoirs of pathogens, survival of mo on surfaces, source of contamination), as part of an epidemiologic investigation, or QA purposes• Media (nutrient‐rich such as TSA or BHI), reagents, and equipment required for surface sampling available in micro lab• Effective sampling requires moisture

Other Microbiologic Sampling

• Biological indicators• Hemodialysis water‐200/ml, 2000/ml• Infant formula‐hospital prepared• Pharmacy‐hospital prepared• Respiratory therapy• Blood bank water bank‐used to thaw plasma• Endoscopes

Microbiologic Sampling of the EnvironmentConclusions

• Do not conduct random microbiological sampling of air, water, and surfaces• When indicated, conduct microbiologic sampling as part of an epidemiologic investigation • Limit microbiologic sampling for QA to: biological monitoring, dialysis water, or evaluation of IC measures


•Lecture Goals•Microbiologic Sampling•Indications•Methods

Reference• CDC Guidelines for Environmental Infection Control in Health‐Care Facilities, 2003. • Rawlinson S, Ciric L, Cloutman‐Green E. J Hosp Infect 2019;103:363‐374• Boyce J. Environmental Evaluation. https://dph.georgia.gov

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Lecture Goals Microbiologic Sampling of the Environment